Microarray biosensors for biological analysis are well known in the art. Generally, a biochemical substance, such as multiple droplets of a reagent, is deposited onto a substrate to form the microarray. Each droplet of the reagent is preferably in electrical communication with an electrode so as to form an individual biosensor. The droplet of reagent may then be exposed to a biological agent or the like, such that a reaction is caused. With the aid of at least one additional electrode, various factors of the reaction may be measured and/or observed to provide the analysis.
Various techniques may be used to create the microarray biosensors, such as screen printing, ink-jetting, micro-pipetting, and pin-deposition. Each of the techniques have advantages. For instance, screen printing may be preferred when a relatively large deposit of a thick reagent layer is required to control spreading of the reagent composition. “Spreading” refers to the outward flow of a deposit of material due to the material's viscosity. Screen printing also may be preferred due to the high speed at which deposition may occur. If smaller area deposits are desired, ink-jetting, micro-pipetting, or pin-deposition may be preferred. These techniques can more precisely control the volume of a reagent composition deposited onto a substrate.
In contrast to screen printing, ink-jetting and micro-pipetting are non-contact techniques, where only the reagent composition contacts the substrate. While ink-jetting can deposit very small volumes of the reagent composition with little spreading, the method cannot effectively deposit biomolecules due to the small size of the individual droplets sprayed by the print head, approximately 10 to 100 picoliters (pL). Furthermore, biomolecules, especially those of high molecular weight, may lead to clogging of the deposition nozzle. Micro-pipetting can deposit very small volumes of the reagent composition, but spreading is influenced by the flow of the reagent composition liquid over the topography of the substrate. If the substrate has raised areas formed by conductors or other features, the reagent composition may flow from the raised areas into lower areas providing more undesirable spreading from the deposition area. As with ink-jetting, biomolecules in the reagent composition also may clog the pipette.
With pin-deposition, one or more microdeposition pins contact the substrate to transfer the reagent composition. Pin contact with the substrate allows for the transfer of relatively small volumes of reagent composition while reducing spreading of the reagent composition from raised features. Thus, pin-deposition may provide thin reagent layers from small deposition volumes while maintaining liquid control advantages.
Pin-deposition generally starts by lowering multiple pins into a source plate having wells containing a reagent composition. As the pins are dipped into the wells, they are coated with the reagent. The pins are moved above the substrate, and the reagent is transferred from the pins to the substrate during a brief touchdown. Pin-deposition also allows the force with which the pin contacts the substrate to be altered.
Conventional deposition pins generally are quill or solid in design. Quill pins differ from solid pins in that quill pins include a narrow slit at the tip of the pin. This slit acts as a fluid reservoir, which holds the reagent composition before the pin contacts the substrate. Each time a quill pin contacts the substrate, it deposits at least a portion of the reagent composition from the reservoir. The reservoir is generally sized to hold sufficient volume of reagent for multiple depositions.
Solid microdeposition pins generally have small, flat tips that are dipped into the reagent composition before each contact with the substrate. As the reagent composition flows down the exterior sides of the pin, there is no passageway for biomolecules and other high molecular weight constituents to clog. The volume and morphology of the resulting deposition spot may depend on the diameter of the solid pin tip, the surface tension of the reagent composition, and the hydrophobic character of the substrate in relation to the hydrophilicity of the reagent composition. For example, a 0.2 millimeter (mm) diameter pin may produce depositions having a diameter of approximately 400 micrometers (μm). When depositing solution volumes of from 10 picoliters (pL) to 10 nanoliters (nL), the 0.2 mm diameter pins may provide deposition areas having diameters ranging from 100 μm to 1,000 μm. Conventional solid pins having tips that are either round or square are known to produce substantially round deposition areas.
However the biosensors are manufactured, they may be used for analysis of a biological fluid, such as whole blood, urine, or saliva. Typically, biosensors have a measurement device that analyzes a sample of the biological fluid placed on a sensor strip. The analysis may determine the concentration of one or more analytes, such as alcohol, glucose, uric acid, lactate, cholesterol, or bilirubin, in the biological fluid. The analysis is useful in the diagnosis and treatment of physiological abnormalities. An ongoing need remains for microdeposition techniques that provide enhanced control over the spreading of the deposited reagent composition.